The isolation of nuclei from tissue-cultured plant cells

A method for the isolation of nuclei from tissue-cultured plant cells is presented. The method employs polyamines as stabilizing agent and uses Percoll for the purification of the nuclei in density gradients. It is applicable to large amounts of tissue and is reasonably quick (500 g of tissue can be processed within 2–3 h). The purified nuclei have retained their morphological characteristics as demonstrated by phase contrast, as well as electron micrographs. High molecular weight DNA can be isolated from these nuclei and histones extracted from the purified nuclei display the expected gel electrophoretic pattern. The retainment of the nucleosomal arrangement is demonstrated by digestion with micrococcal nuclease and subsequent analysis of the DNA fragments on agarose gels. The purified nuclei contain high activities of several nucleus-specific enzymes such as α-amanitin-sensitive and insensitive RNA polymerases, protein kinases, poly-ADP-ribosylating enzymes and DNA polymerases.